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Background and Purpose: Cerebrospinal fluid (CSF) real-time quaking-induced conversion (RT-QuIC) has a high degree of sensitivity and specificity for the diagnosis of sporadic Creutzfeldt-Jakob disease (sCJD) and this has led to its being included in revised European CJD Surveillance Network diagnostic criteria for sCJD. As CSF RT-QuIC becomes more widely established, it is crucial that the analytical performance of individual laboratories is consistent. The aim of this ring-trial was to ascertain the degree of concordance between European countries undertaking CSF RT-QuIC., Methods: Ten identical CSF samples, seven from probable or neuropathologically confirmed sCJD and three from non-CJD cases, were sent to 13 laboratories from 11 countries for RT-QuIC analysis. A range of instrumentation and different recombinant prion protein substrates were used. Each laboratory analysed the CSF samples blinded to the diagnosis and reported the results as positive or negative., Results: All 13 laboratories correctly identified five of the seven sCJD cases and the remaining two sCJD cases were identified by 92% of laboratories. Of the two sCJD cases that were not identified by all laboratories, one had a disease duration >26 months with a negative 14-3-3, whilst the remaining case had a 4-month disease duration and a positive 14-3-3. A single false positive CSF RT-QuIC result was observed in this study., Conclusions: This study shows that CSF RT-QuIC demonstrates an excellent concordance between centres, even when using a variety of instrumentation, recombinant prion protein substrates and CSF volumes. The adoption of CSF RT-QuIC by all CJD surveillance centres is recommended., (© 2022 The Authors. European Journal of Neurology published by John Wiley & Sons Ltd on behalf of European Academy of Neurology.)

BK-virus (BKV) associated nephropathy (BKVAN) and BKV associated haemorrhagic cystitis (HC) are complications of BKV infection/reactivation in renal and allogeneic haematopoietic stem cell transplantation (HSCT) patients, respectively. The task of how to manage these diseases was given to the chair by the Swedish Reference Group for Antiviral Therapy (RAV). After individual contributions by members of the working group, consensus discussions were held in a meeting on 23 January 2018 arranged by RAV. Thereafter, the recommendations were published in Swedish on November 2018. The current translation to English has been approved by all co-authors. High BKV serum levels suggest an increased risk for BKVAN and potential graft failure. For detection of BKVAN, careful monitoring of BKV DNA levels in serum or plasma is recommended the first year after renal transplantation and when increased creatinine serum levels of unknown cause are observed. Notably, a renal biopsy is mandatory for diagnosis. To reduce the risk for progression of BKVAN, there is no specific treatment, and tailored individual decrease of immunosuppression is recommended. For BKV-HC, BKV monitoring is not recommended, since BK-viruria frequently occurs in HSCT patients and the predictive value of BKV in plasma/serum has not been determined. However, the risk for BKV-HC is higher for patients undergoing myeloablative conditioning, having an unrelated, HLA-mismatched, or a cord blood donor, and awareness of the increased risk and early intervention may benefit the patients. Also for BKV-HC, no specific therapy is available. Symptomatic treatment, e.g. forced diuresis and analgesics could be of use.

Introduction: To study the presence of European bat lyssavirus (EBLV) infections in bat reservoirs in Sweden, active surveillance was performed during the summers from 2008 to 2013., Material and Methods: Bat specimens were collected at >20 bat colonies in the central, southeastern, and southern parts of Sweden. In total, blood and saliva of 452 bats were examined by a virus neutralization test and by reverse transcription polymerase chain reactions (RT-PCRs)., Results and Discussion: EBLV neutralizing antibodies were detected in 14 Daubenton's bats ( Myotis daubentonii ), all trapped in Skåne or Småland (south and southeast of Sweden). The result was not unexpected since EBLV has been shown to be present in many neighboring countries, for example, Denmark, Finland, Germany, and Norway. However, Sweden has been regarded free of rabies in terrestrial mammals since 1896. Although very rare, spillover of EBLV into other animals and humans have occurred, and the risk of EBLV infection to other species including humans should not be ignored. This is the first report of lyssavirus infection in Swedish bats.

Molluscum contagiosum is a viral infection of the epidermis characterized by skin-colored papules or nodules frequently with a central depression. Atypical variants may occur, primarily in immunosuppressed individuals. We here report a case of »giant Molluscum contagiosum« in an immunocompetent child. The patient was presented with a fairly smooth nodule of 2 cm in diameter on the ring finger. Molluscipoxvirus-like virus particles were detected by electron microscopy from the nodule, but since the clinical picture was not compatible with MC, next generation sequencing was performed in order to verify the diagnosis.  Of the total number of obtained sequences, 25% belonged to molluscipoxvirus (MCV) and de novo assembly revealed three contigs corresponding to 95% of the MCV genome. The assembled genome was compared to previously published sequences of the »major envelope protein« used for genotyping of MCV genus. Several unique single nucleotide polymorphisms were identified, which led us to classify this virus as a new subtype of MCV.

At present, the testing of 14-3-3 protein in cerebrospinal fluid (CSF) is a standard biomarker test in suspected sporadic Creutzfeldt-Jakob disease (sCJD) diagnosis. Increasing 14-3-3 test referrals in CJD reference laboratories in the last years have led to an urgent need to improve established 14-3-3 test methods. The main result of our study was the validation of a commercially available 14-3-3 ELISA next to the commonly used Western blot method as a high-throughput screening test. Hereby, 14-3-3 protein expression was quantitatively analyzed in CSF of 231 sCJD and 2035 control patients. We obtained excellent sensitivity/specificity values of 88 and 96% that are comparable to the established Western blot method. Since standard protocols and preanalytical sample handling have become more important in routine diagnostic, we investigated in a further step the reproducibility and stability of 14-3-3 as a biomarker for human prion diseases. Ring trial data from 2009 to 2013 revealed an increase of Fleiss' kappa from 0.51 to 0.68 indicating an improving reliability of 14-3-3 protein detection. The stability of 14-3-3 protein under short-term and long-term storage conditions at various temperatures and after repeated freezing/thawing cycles was confirmed. Contamination of CSF samples with blood appears likely to be an important factor at a concentration of more than 2500 erythrocytes/μL. Hemolysis of erythrocytes with significant release of 14-3-3 protein started after 2 days at room temperature. We first define clear standards for the sample handling, short- and long-term storage of CSF samples as well as the handling of blood- contaminated samples which may result in artificially elevated CSF levels of 14-3-3.

Objective: The objective of this report is to assess the presence and viral load of JC polyomavirus (JCV) DNA in cerebrospinal fluid (CSF) and plasma from neuropsychiatric systemic lupus erythematosus (NPSLE) patients in comparison to controls and to investigate if different types of immunosuppressive treatments were correlated to detection and viral load of JCV DNA in SLE., Background: Reactivation of a latent JCV infection with subsequent development of the fatal disease progressive multifocal leukoencephalopathy (PML) has become an increasing problem in patients with autoimmune diseases treated with newer immunosuppressants. Accumulating data point out that SLE patients are at particular risk for PML compared to patients with other rheumatic diseases., Methods: CSF samples (n = 69) and plasma samples (n = 51) from 71 SLE patients and 58 controls (53 CSF samples and 50 plasma samples) with other non-inflammatory neurological disease (OND) were analyzed for JCV DNA with a quantitative PCR method., Results: All CSF and plasma samples from NPSLE patients and controls were negative for JCV DNA., Conclusion: JCV DNA was absent in CSF and plasma in NPSLE patients and controls and consequently we were not able to identify any correlation between the occurrence of JCV DNA and type of immunosuppressive medication.

BK virus associated nephropathy occurs in approximately 5% of renal transplant recipients. Quantitation of BKV DNA in serum/plasma early in the course of disease has been suggested to be an important diagnostic tool for polymavirus-associated nephropathy (PVAN). The aim of this study was to develop a BKV real-time PCR (qPCR), which could be included in a diagnostic qPCR platform. Additionally, the significance of the assay as a surrogate marker for PVAN was investigated. Quantitation of BKV DNA by qPCR was carried out on 234 serum samples from a retrospective study including 31 renal transplant recipients monitored for at least 6 months post-transplantation. BKV viremia was detected in 9 out of 31 patients. Four patients had a viral load of >10,000 copies/ml at least on one occasion. In two of these patients, PVAN was diagnosed clinically during the study period. In retrospect, these patients were shown to be BKV positive before the clinical diagnosis of PVAN was made. Another two patients had a permanent graft dysfunction, but were never clinically diagnosed with PVAN. None of the remaining five patients with BKV DNA (<10,000 copies/ml) had renal impairment. Based on these results, an algorithm was introduced at the study center in 2006 and to date, August 2011, no cases of PVAN with loss of graft have been observed. The concept of including different PCR protocols in a common qPCR platform allows laboratories with small sample numbers to perform regularly a variety of assays at a reasonable cost., (Copyright © 2011 Wiley-Liss, Inc.)

We describe a patient with facial cellulitis/erysipelas due to cowpox virus inoculation in the respiratory epithelium of the nose. A cytopathic agent was isolated in cell culture, and the diagnosis of cowpox was confirmed by electron microscopy and polymerase chain reaction. The most likely source of infection was exposure to the family cats. In addition to the severe edematous cellulitis of the face, the clinical course was dominated by several areas of subcutaneous, necrotizing lymphadenitis, from one of which a huge abscess formed that had to be incised. Hyperbaric oxygen treatment was provided to prevent development of dermal necrosis. The healing process in the numerous areas of lymphadenitis was markedly protracted, and 1 persisting node (which yielded positive results on polymerase chain reaction) had to be excised 2 years after onset of disease. This is the first reported case of inoculation of cowpox virus in the respiratory mucosa of the nose. It resulted in a clinical course totally different than that for inoculation in the skin. We also present a short review of findings on orthopoxvirus infection that focuses on the chain of transmission.

The human polyomaviruses BK (BKV) and JC (JCV) affect immunosuppressed patients and are associated with urogenital tract (BKV) and CNS disorders (JCV) and in humans, the pathogenic role of the rhesus monkey virus, Simian virus 40 (SV40), is uncertain. These three viruses have somewhat overlapping tissue pathogenicity and detection of all three polyomaviruses is desirable. A broadly targeted, simple, single tube real-time degenerated quantitative PCR (QPCR) technique for detection of JCV, BKV and SV40 DNA was developed. To avoid false positive results, due to contamination with commonly used SV40 T-antigen plasmids, a conserved region of the VP2 gene was targeted. Down to 1-10 copies of target DNA per PCR reaction were detected. The QPCR was compared with a nested PCR on 41 clinical samples (urine, serum and plasma): 24 (58.5%) tested positive by nested PCR, whereas 31 (75.6%) were positive with QPCR. One CSF sample, from a patient with progressive multifocal leukoencephalopathy, was negative with the nested PCR but determined as positive by QPCR. Sera from 24 blood donors were negative with QPCR. The QPCR described had a high sensitivity. Its specificity was confirmed sequencing. The QPCR is simple to perform and is valuable for diagnosis of polyomavirus infection.

Rabies has become a disease of increasing concern. One reason is that bat variant rabies is a more common cause of human disease, with 1-2 deaths per year in the United States. Bat bites are much more difficult to document than bites from larger animals. Deaths from rabies encephalitis have remained undiagnosed until postmortem examination. Prophylaxis includes a series of 5 vaccinations during 28 days. Vaccine efficacy has been documented, even in young children.

Creutzfeldt-Jakob disease (CJD) is a prion disease characterized by rapid neurodegeneration that leads to the death of the patient within months to a few years. Since the disease is transmissible, there is an obligation in Sweden to report possible CJD cases to the Swedish Institute for Infectious Disease Control. To make a diagnosis of CJD is difficult, especially early in the course of the disease when the clinical features may be very vague and heterogeneous. Hence, accurate biological markers both for confirming and excluding CJD would be of great value. The currently recommended investigation of a patient with possible CJD comprises clinical evaluation. electroencephalography, computed tomography or magnetic resonance imaging of the brain and test for 14-3-3 protein in the cerebrospinal fluid (CSF). Recent studies suggest that analysis of total tau (T-tau) and phospho-tau (P-tau) in CSF is a valuable complement to this set of investigations. Here, we review how CSF T-tau and P-tau may aid in the diagnosis of CJD and illustrate this by presenting cases from routine clinical practice.

Vaccination against smallpox is again considered in order to face a possible bioterrorist threat, but the nature and the level of the immune response needed to protect a person from smallpox after vaccination are not totally understood. Therefore, simple, rapid, and accurate assays to evaluate the immune response to vaccinia virus need to be developed. Neutralization assays are usually considered good predictors of vaccine efficacy and more informative with regard to protection than binding assays. Currently, the presence of neutralizing antibodies to vaccinia virus is measured using a plaque reduction neutralization test, but this method is time-consuming and labor-intensive and has a subjective readout. Here, we describe an innovative neutralization assay based on a modified vaccinia virus Ankara (MVA) vector expressing the green fluorescent protein (MVA-gfp). This MVA-gfp neutralization assay is rapid and sensitive and has a high-throughput potential. Thus, it is suitable to monitor the immune response and eventually the efficacy of a large campaign of vaccination against smallpox and to study the vector-specific immune response in clinical trials that use genetically engineered vaccinia viruses. Most importantly, application of the highly attenuated MVA eliminates the safety concern in using the replication-competent vaccinia virus in the standard clinical laboratory.

Human polyomavirus BK (BKV; GenBank or EMBL or DDBJ accession no. NC001538) is often reactivated in immunosuppressed patients. Reactivation has been associated primarily with excretion of the virus in the urine, and there have been few reports of renal and/or neurological disease caused by BKV in patients with acquired immunodeficiency syndrome (AIDS). Polymerase chain reaction, Southern blotting, and sequencing were used to detect and identify the noncoding control region (NCCR) of BKV in different tissues in an AIDS patient with meningoencephalitis, retinitis, and nephritis. An undescribed reorganized NCCR variant of the virus, completely different from the variants detected in peripheral blood leukocytes (PBLs) and urine, was identified in the cerebrospinal fluid (CSF) and CNS tissues. These results suggest that rearrangements in the NCCR of the virus have resulted in a BKV variant, which is better adapted to the host cell machinery of the cells in CNS tissue. The rearranged variant (BKV CNS) might have been involved in the initiation and/or development of the pathological lesions observed in the CNS-related tissues of this patient., (Copyright 2003 Wiley-Liss, Inc.)

In June 2000, a case of rabies was diagnosed in Stockholm. The patient, a 19-year-old woman, had been bitten by a dog in Thailand three months earlier. She was admitted with a 2-day history of pain and paresthesia at the exposure site (right arm), along with anxiety. Her neurological symptoms progressed, and during the following week she developed the typical signs of furious rabies. Despite intensive care, her condition deteriorated continuously, and she died 18 days after onset of symptoms. The diagnosis was not considered until five days after admission to the hospital. A saliva sample was obtained and the diagnosis confirmed by virus isolation in mouse neuroblastoma cells. Although Sweden is free of rabies, the diagnosis should be considered in patients with encephalitis after having visited a rabies endemic area. Tourists must be informed of the vital importance of post-exposure prophylaxis after suspected infection.

Background: The two widely spread human polyomaviruses, BK virus (BKV) and JC virus (JCV) establish latency in the urinary tract, and can be reactivated in AIDS. JCV might cause progressive multifocal leucoencephalopathy, but although up to 60% of AIDS patients excrete BKV in the urine there have been few reports of BKV-related renal and/or neurological disease in AIDS., Objective: To report on an AIDS patient with progressive renal and neurological symptoms involving the retina., Design: Case report., Setting: Venhälsan, Söder Hospital, Stockholm, Sweden., Methods: The brain, eye tissue, cerebrospinal fluid, urine and peripheral blood mononuclear cells were analysed by nested PCR for polyoma-virus DNA. Macroscopical and microscopical examination were performed of the kidney and brain post mortem. Immunohistochemical stainings for the two BKV proteins, the VP1 and the agnoprotein, were performed on autopsy material and virus infected tissue culture cells., Results: BKV could be demonstrated in the brain, cerebrospinal fluid, eye tissues, kidneys and peripheral blood mononuclear cells., Conclusion: During 6 years, approximately 400 cerebrospinal fluid samples from immunosuppressed individuals with neurological symptoms have been investigated by PCR for the presence of polyomaviruses. BKV DNA has, so far, only been found in the case reported here. Although reports of BKV infections in the nervous system are rare, there is now evidence for its occurrence in immunocompromised patients and the diagnosis should be considered in such patients with neurological symptoms and signs of renal disease. The diagnosis is simple to verify and is important to establish.

Objective: To report the clinical and histopathologic characteristics of BK virus (BKV) retinitis., Design: Case report., Testing: The clinical features of bilateral retinitis in a 29-year-old homosexual white male with the acquired immune deficiency syndrome (AIDS) included focal, mottled fundus pigmentation, and haloes, as documented by fundus photography. After death of the patient, the left eye was studied by light microscopic and immunohistochemical examination. The nested polymerase chain reaction (PCR) was used to detect viral deoxyribonucleic acid (DNA) in the right eye and other nonocular tissues. The specificity was then confirmed by restriction enzyme analysis., Results: The retina of the left eye showed focal necrosis and contained cells with intranuclear staining for the BKV VP1 protein. In the right eye, BKV DNA was detected in the retina and other tissues by nested PCR. Autopsy showed that BKV infection was also present in the brain, kidneys, and peripheral blood mononuclear cells., Conclusions: A number of pathogens may cause retinitis in patients with AIDS. The authors have shown that BKV should be included among those pathogens and that some clinical features may suggest the presence of BKV retinitis.

JC virus (JCV) DNA was detected in cerebrospinal fluid (CSF) samples from patients with progressive multifocal leukoencephalopathy (PML) but not in CSF samples from patients with herpes simplex encephalitis, enteroviral meningitis, or multiple sclerosis. This suggests that inflammatory processes in the brain do not necessarily reactivate JCV, which further supports the proposal that the presence of JCV DNA in the CSF is diagnostic for PML.

The sensitivity and specificity of PCR of CSF for the diagnosis of progressive multifocal leuko encephalopathy is estimated at 75 and 98.5%, respectively. However, inter-laboratory and inter-technique variations have been shown to produce wide variations. A 10-fold dilution series of JC virus in cerebrospinal fluid was prepared and circulated for 'blind' evaluation in laboratories participating in a European Union Concerted Action on Virus Meningitis and Encephalitis. Six of seven laboratories returned results with sensitivity of between 10 and 1 JCV DNA copy equivalents per 10 microl of CSF, one laboratory detected 10(5) copies per 10 microl of CSF. These results demonstrate the feasibility of using virus diluted in CSF for comparison of PCR techniques, and that the range of sensitivity of JCV PCR in proficient laboratories is between 10 and 1 copy equivalents per 10 microl of CSF.

Two polyomaviruses, JC virus (JCV) and BK virus (BKV), affect humans. JCV is the causative agent of progressive multifocal leukoencephalopathy (PML), and detection of JCV in the central nervous system (CNS) is a prerequisite for confirmation of the disease. BKV is generally not associated with neurological disease, but involvement of BKV in patients with CNS disorders has been reported. In the present study polyomavirus DNA was detected by a nested PCR at a sensitivity corresponding to the detection of 10 copies of JCV DNA in 10 microliters of cerebrospinal fluid (CSF). CSF samples from 212 patients with neurological symptoms and immunodeficiencies were investigated for the presence of polyomavirus DNA. Of 128 human immunodeficiency virus (HIV)-infected patients, 14 (11%) had JCV DNA in their CSF, and all 14 patients had clinical PML. BKV DNA was detected in one HIV-infected patient with neurological symptoms not compatible with PML. Among 84 HIV-negative patients, 6 (7%) had detectable JCV DNA in their CSF, confirming PML in patients with clinical conditions of T-cell lymphoma, chronic lymphatic leukemia, status postliver transplantation, congenital immunodeficiency, sarcoidosis, and immunodeficiency of unknown origin. The specificity of the PCR was confirmed by a clinical follow-up study which showed full agreement between the detection of JCV DNA in CSF and clinically manifest PML. The described PCR is a rapid, reproducible, and easily performed assay.

Objectives: To investigate if JC virus (JCV) can be involved in the pathogenesis of Alzheimer's disease (AD) and astrocytomas., Study Design: A nested polymerase chain reaction (PCR) was used for the detection of JCV DNA in autopsy brain material (cerebral white matter) and cerebrospinal fluid (CSF) specimens from patients with AD and age-matched control patients without neurological diseases, together with biopsies from patients with astrocytomas (grades 3 and 4). Brain autopsy material from AIDS patients with progressive multifocal leukoencephalopathy (PML) was examined as positive control material., Results: JCV DNA was detected by PCR in only one of the 17 brain autopsies from patients with AD, but in none of the 26 control patients without neurological diseases and in none of the 5 astrocytoma biopsies. JCV DNA was, however, detected in the brain material from two patients with PML., Conclusion: Our results show that JCV infection does not seem to be directly involved in the pathology of AD or in the development of astrocytomas. In addition, since no viral DNA was detected in CSF specimens from 43 patients without PML (17 with AD and 26 elderly controls), our results suggest that the finding of JCV DNA in CSF correlates to PML.

Background: A nested polymerase chain reaction (PCR) was developed to detect BK virus (BKV) and JC virus (JCV) DNA sequences. The unique clevage site for BamHI restriction enzyme was located in the JCV amplimer and cleavage was used to differentiate between BKV and JCV., Study Design: Twenty-three urine specimens from 17 bone marrow recipients with haemorrhagic cystitis and one liver transplant patient were tested for the presence of BKV and JCV DNA. Four brain tissue specimens (paraffin embedded brain tissues and a fresh frozen brain biopsy) and 5 cerebrospinal fluids from 3 AIDS patients and one liver transplant patient, all with progressive multifocal leukoencephalopathy (PML), were also examined by PCR., Results: The sensitivity of the PCR was 10 genomes for each virus. BKV DNA was detected in 15 urine specimens from 12 bone marrow transplant patients. JCV DNA was detected in 4 cerebrospinal fluids and 4 brain tissues from patients with PML., Conclusion: Our results show that the nested PCR is a sensitive and rapid assay that can be used for diagnosis of BKV and JCV infections. The cerebrospinal fluid appears to be a suitable material for diagnosis of JC virus reactivation in the brain.